Many natural amino acids are prepared in enantiomerically pure form by fermentation using genetically modified bacteria (de Graaf, et al., Adv. Biochem. Eng. Biotechnol. 73:9–29 (2001); Sahm, et al., Ann. NY Acad. Sci. 782:25–39 (1996)). Although not all proteinogenic amino acids (and only very few unnatural or D-amino acids) can be prepared in this way, chemical syntheses for enantiomerically pure amino acids are very costly. As a result, several enzymatic processes have been developed, and used on a scale of several metric tons per year. The methods range from kinetic racemate cleavage with the aid of acylases, amidases, esterases, hydantoinases, amino acid oxidases and proteases, to enantioselective syntheses by means of lyases, aminotransferases and dehydrogenases (Schmid, et al., Curr. Opin. Biotechnol. 13(4):359–366 (2002)).
In addition to enantioselective syntheses, enantiomerically enriched amino acid preparations may be obtained by dynamic kinetic racemate cleavages, in which the unwanted enantiomer is racemized in situ. A 100% yield can be achieved by combining a D- or L-amino acid oxidase with an unselective chemical reduction of the incipient imino acid back to the principal amino acid. The reducing agent, e.g., NaBH4, must, however, be employed in an excess of at least 25 equivalents, which makes this option very costly (Enright et al., Chemical Communications 20:2636–2637 (2003)).
The amination of α-ketonic acids by amino acid dehydrogenases is generally known. Although the educt is many times more costly than, for example, the corresponding racemic amino acid, by coupling an amino acid oxidase with an amino acid dehydrogenase the corresponding ketonic acid can be obtained in situ from an amino acid. When both of the enzymes have the opposite enantio-selectivity, a D-amino acid can be completely converted into an L-amino acid or an L-amino acid converted into a D-amino acid. Thus, starting with a racemate, an enantiomerically pure compound can be produced. In order to make the process economically viable, the NADH cosubstrate must be enzymatically regenerated. Enzymes such as formate dehydrogenase and malate dehydrogenase (decarboxylating) which liberate carbon dioxide from its substrate and thus make the reaction irreversible, are particularly well suited for this (Hanson, et al., Bioorganic Medicinal Chem. 7(10):2247–2252 (1999); Nakajima, et al., J. Chem. Soc. Chem. Commun. 13:947–8 (1990)).

For a rapid and complete conversion in the cell-free system, a catalase must also be present. This is needed because hydrogen peroxide is produced in the oxidative step catalyzed by the amino acid oxidase and this leads to the decarboxylation of the ketonic acid and to deactivation of enzymes (Trost, et al, J. Mol. Catalysis B: Enzymatic 19–20:189–195 (2002)). The conversion of racemate into enantiomerically pure amino acids is possible in this system with >99% ee and >95% yield. However, this method is costly, since four different enzymes have to be separately prepared and isolated.